Prostate-specific membrane antigen (PSMA) is among the most intensively targeted biomarkers for imaging metastatic prostate cancer. PSMA is a zinc-dependent metallopeptidase that catalyzes the hydrolysis of a series of N-acylpolygammaglutamate derivatives (Mesters, 2006; Barinka, 2004; Pinto, 1996). It is expressed within certain normal tissues, (Ghosh, 2004) but transitions to abundant plasma membrane expression in the epithelium of most prostate cancer and within other solid tumor neovasculature (Rajasekaran, 2005). PSMA membrane expression is associated with metastasis (Chang, 2001), castration resistance (Bander, 2005) and progression of prostate cancer (Perner, 2007).
Several different scaffolds are available for synthesis of small-molecule PSMA inhibitors, and have been reviewed (Byun, 2009; Tsukamoto, 2007; Zhou, 2005). As shown FIG. 1, those potent scaffolds share common features, namely; a) a pentanedioic acid as a glutamate mimic to fit within the 51′ binding pocket of the PSMA active site; and b) a zinc-binding group to interact with the catalytic zinc atom at the PSMA active site. A substituent (R) can reside either within the 51 binding pocket or within a void in the protein that extends to the surface. Scaffolds composed of phosphonates or phosphinates, (Jackson, 1996; Jackson, 2000), phosphoramidates (Maung, 2004) and ureas (Kozikowski, 2001; Kozikowski, 2004; Maung, 2004) of general structures 1-3, as well as thiol 4 (Majer, 2003; Stoermer, 2012) and hydroxamate 5 (Stoermer, 2003) have been reported as effective zinc binding groups for PSMA inhibition. However, the presence of a zinc binding moiety and a glutamate mimic residing in the S1′ pocket are themselves not sufficient for high binding as demonstrated by gly-urea-glu compound 8 (FIG. 2A and FIG. 2B) (Wang, 2013). Of the reported PSMA binding scaffolds, urea-based inhibitors, first introduced by Kozikowski in 2001 (Kozikowski, 2001) for inhibition of glutamate carboxypeptidase II within the central nervous system, have been utilized the most for targeting PSMA due to their high binding affinity and synthetic simplicity (Kozikowski, 2001; Kozikowski, 2001; Foss, 2012; Mease, 2013; Vargas, 2015; Chen, 200; Barinka, 2008). A variety of low-molecular-weight compounds based on the various scaffolds discussed above, primarily the ureas, have been labeled with radionuclides for positron emission tomography (PET) and single photon emission computed tomography (SPECT), namely, 125/124I, 99mTc, 111In, 18F, 11C, 68Ga, 64Cu, and 86Y, and have demonstrated PSMA-targeted imaging of prostate cancer in experimental models (Foss, 2012; Mease, 2013; Vargas, 2015; Banerjee, 2008; Banerjee, 2015; Banerjee, 2010; Banerjee, 2014; Chen, 2008; Chen, 2011; Eder, 2012; Foss, 2005; Hillier, 2013; Kularatne, 2009; Lapi, 2009; Maresca, 2009; Mease, 2008; Ray Banerjee, 2013; Weineisen, 2014). Several of these have been translated to phase 0-1 clinical trials, where they have enabled visualization of both primary and metastatic bone and soft-tissue lesions due to prostate cancer. (Afshar-Oromieh, 2015; Afshar-Oromieh, 2012; Afshar-Oromieh, 2013; Afshar-Oromieh, 2014; Barrett, 2013; Cho, 2012; Rowe, 2015; Szabo, 2015; Vallabhajosula, 2014). In addition, one agent [18F]DCFPyL has also shown a higher sensitivity for detection of metastatic clear cell renal carcinoma compared to conventional imaging methods (S. P. Rowe, 2015).
However, clinical imaging studies also exhibited considerable uptake in non-target PSMA-expressing tissues such as the salivary glands and kidneys, bringing to light potential dose-limiting off-target effects, particularly for radiotherapeutic analogs. Additional PSMA-binding scaffolds that might preserve the positive imaging characteristics of the ureido scaffolds but clear from the non-target organs were sought. The carbamate scaffold was chosen because it would retain the overall geometry of the existing inhibitors, differing only with an O for NH substitution, which eliminates a potential hydrogen bonding donor group present in the ureas. The only PSMA-binding carbamate reported is gly-amino-pentanedioic acid 1 (Wang, 2013) displayed low binding affinity to PSMA, most likely due to the absence of productive binding within the S1 pocket, similar to ureido compound 8. Herein a new class of potent PSMA inhibitors based on the carbamate scaffold to maintain glutamate and S1 pocket side chain geometry and for putative binding to zinc have been reported. Carbamate scaffolds may complement the existing urea and other scaffolds upon which inhibitors, imaging and therapeutic agents targeting PSMA have been based.
Because of the favorable pharmacokinetic profile of this class of compounds, i.e., low nonspecific binding, lack of metabolism in vivo and reasonable tumor residence times, it has been reasoned that urea and carbamate-based agents could also be used for molecular radiotherapy. This will be in analogy with radioimmunotherapy (RIT), which has proved remarkably successful in the treatment of lymphoma with two commercial products routinely integrated into clinical practice. However, (RIT) is fraught with difficulties inherent in the use of radiolabeled antibodies for imaging, including prolonged circulation times, unpredictable biological effects and the occasional need for pre-targeting strategies. Furthermore, antibodies may have less access to tumor than low molecular weight agents, which can be manipulated pharmacologically. Therefore a need remains for low molecular weight compounds with high binding affinity to PSMA for the imaging and radiotherapy of tumors.
Further, for the aforementioned reasons, fluorescent-linker-carbamate based PSMA inhibitors have also been investigated. Targeted fluorescent PSMA binding compounds may find utility in fluorescence guided surgery and biopsy of PSMA positive tumors and tissues, the former providing visual confirmation of complete removal of PSMA containing tissue. Moreover, carbamate conjugates of photosensitizing dyes also provide PSMA targeted photodynamic therapy agents.